The overall goal of this work is to develop anatomical, functional, and molecular magnetic resonance imaging (MRI) techniques that allow non-invasive assessment of brain function and apply these tools to study plasticity and learning in the rodent brain. MRI techniques are having a broad impact on understanding brain. Anatomical based MRI has been very useful for separating gray and white matter and detecting numerous brain disorders. Functional MRI techniques enable detection of regions of the brain that are active during a task. Molecular MRI is an emerging area, whose major goal is to image a large variety of processes in tissues. The goal of this project is to translate MRI developments in all these areas to study system level changes that occur in the rodent brain during plasticity and learning. Aim 1: Over the past few years, we have completed studies in the rodent brain that acquired very high temporal and spatial resolution functional MRI (fMRI) to monitor changes in hemodynamics as a surrogate marker of electrical activity during forepaw stimulation. Work over the past year has focused on trying to separate macrovascular contribution (draining venuoles) from microvascular contributions (arterioles/capillaries)in mid cortex. The general thinking is that if fMRI maps can be made from microvascular contributions this will more accurately reflect neural function. High spatial and temporal resolution fMRI indicated that significant activation could be detected prior to venuoles contributing in MRI of the whisker barrel cortex. This represents one of the first studies that have directly detected the different contributions of the vascular network to BOLD fMRI. Aim 2: Over the past several years we have demonstrated that manganese chloride enables MRI contrast that defines neural architecture, can monitor activity, and can be used to trace neural connections. Over the last couple of years we have completed the assignment of cortical layers detected using manganese enhanced MRI by comparison to histology and have demonstrated that functional anatomy of several cortical regions of the rodent brain can be defined in individual animals. In particular, clear cytoarchitectural boundaries can be detected between numerous brain areas enabling, for the first time, cytoarchitectural changes to be followed in individual brains over time. Over the past year we have been developing computational techniques that enable automatic and unbiased extraction of anatomical boundaries from these images. This is of growing importance because of recent work from the Duyn group that demonstrates that cytoarchitectural information can be obtained from human MRI. In addition, we have completed studies that trace the laminar inputs of the olfactory pathway from the olfactory bulb to rodent frontal cortex. The anatomic projections from olfactory cortex to frontal cortex have not previously been measured. The manganese based MRI predictions have been confirmed by classical histological based neural tracing techniques. Experiments to assess whether manganese enhanced MRI can detect changes in connections due to olfactory learning and genetic effects in olfactory learning have begun. Aim 3: Functional MRI studies were performed to measure changes in brain activation that occur after denervation of peripheral nerves. Previously we have studied the plasticity that occurs after denervation of the forepaw and hindpaw usign fMRI and demonstrated using manganese enhanced MRI that there were laminar specific changes in cortical inputs and outputs. In order to perform physiological experiments to assess the mechanism for these changes we decided to switch to studying the whisker barrel cortex where much more is known about detailed electrophysiology. Similar to the forepaw and hindpaw, denervation causes plasticity which leads to increased activation along the unaffected pathway and ipsilateral activation in the affected cortex. fMRI and manganese enhanced MRI showed a strengthening of thalalmo-cortical inputs on the unaffected pathway that have been verified in slice electrophysiological studies. This is quite interesting since it is widely believed that in the adult barrel cortex this synpase can not be strengthened. Future work will address the mechanisms for this strenghtening as well as begin to study the basis of the ipsilateral activation detected. Aim 4: We have begun to explore the use of advanced MRI tools for studying simple learning paradigms in the rodent. In order to accomplish this we have been developing techniques that will enable routine fMRI in awake rodents. While fMRI is widely performed in humans and awake primates there have only been a few scattered studies on awake rodents. Training regimens and techniques to hold the head have been developed and intial fMRI results are being obtained. Over the past year we have been able to get consistent fMRI responses due to visual and somatosensory stimulation. The awake rodent fMRI protocol is being used to study brain changes during olfactory induced fear conditioning.